Estrogens are major regulators of many physiological functions, such as those associated with reproduction and mammary gland development (George et al. The physiology of Reproduction, 1988; Vol. I, page 3., Raven Press, New York). Estrogens influence growth, differentiation, and function of many target tissues including tissues of female and male reproductive tract. Estrogens also play an important role in the maintenance of bone mass and in the cardiovascular system where estrogens have certain protective effects (Farhat et al. FASEB J. 1996; 10:615-624). The effects of estrogen are mediated in either normal cell or neoplastic target cells via an initial interaction with the estrogen receptor. Estrogen receptors (alpha (ERα) and beta (ERβ)) belong to a nuclear hormone receptor superfamily of transcription factors (Green et al. Nature 1986;320:134-139, Kuiper et al. Proc. Natl. Acad. Sci. USA. 1996;93(12):5925-30, Mosselman et al. FEBS Lett 1996 Aug. 19;392(1):49-53, Enmark et al. J. Clin. Endocrinol. Metab. 1997;82(12):4258-65, Bhat et al. J Steroid Biochem Mol Biol 1998;67(3):233-40). These receptors play a critical role in hormonal modulation of gene expression by estrogen and estrogen-like ligands. Signal transduction upon ligand-binding is dependent on characteristic sequence motifs within the receptor protein. These may include a DNA binding domain (DBD), nuclear localization signals, a ligand-binding domain (LBD) and transactivation domains, TAF-1 and TAF-2 which activate transcription of target genes in a ligand independent or dependent fashion respectively. Estrogen receptors dimerize upon ligand activation and this process precipitates DNA binding (Cowley et al. J Biol Chem 1997 Aug. 8;272(32):19858-62, Pettersson et al. Mol. Endocrinol. 1997; Sep. 11(10):1486-96, Pace et al. J. Biol. Chem. 1997;272(41):2,5832-8). Ligand-bound receptor recognizes specific estrogen response elements within the promoter regions of estrogen-regulated genes to induce the transactivation response.
It has been shown that members of the estrogen receptor (ER) superfamily have multiple subtypes and isoforms. ER-like mRNAs distinct from the wild-type ER mRNA have been identified in many known ER positive tissues and cell lines. These isoforms have been mostly derived from alternative splicing of ER mRNAs (Lu et al. Mol Cell Endocrinol. 1998 Mar. 16;138 (1-2):199-203, (Murphy et al., J. Steroid Biochem. Mol. Bio. 1998; 65:175-180, Lu et al., Mol. Cell. Endrocrinol. 1998; 16: 199-203, Murphy et al., J. Steroid Biochem. Mol. Bio. 1997; 62: 363-372). Furthermore, there has been strong evidence that some variant/mutant ER mRNAs are stably translated in vivo and that they may have functional role(s) possibly in ER signal transduction (Fuqua et al., Cancer Research, 1992; 52, 483-486, Fuqua et al., Cancer Research, 1991; 51, 105-109). Recently several isoforms of the human ERβ gene have been described (WO 99/07847, Kastner et al. Proc. Natl. Acad. Sci. USA, 1997; 87, 2700-2704, Leroy et al. EMBO J. 10, 59-69, 1991, Zelent et al. EMBO J. 10, 71-81, 1991). Accordingly, these variant ER mRNAs produce novel proteins which differ structurally and exhibit altered physiological functions. For example, some of ER variant receptors possess anomalous transcriptional activity that may inhibit or enhance the effects of the wild-type receptors. In addition, some may act as dominant negative receptors. In one example, adenoviral delivery of a dominant negative ER to ER-positive breast cancer cells effectively suppressed estrogen-stimulated cell proliferation and the hormonal induction of endogenous genes (Lazennec et al. Mol Endocrinol. 1999; 13(6):969-80.). In another example, five transcripts arise from the human estrogen receptor β (ERβ) (Moore et al. Biochem Biophys Res Commun 1998; 247(1):75-8). In this case, full-length variants showed reduced affinity for estrogens and were able to form DNA-binding homodimers and heterodimers with each other and with the ERβ subtype. In another example, the sequence of a splice variant receptor, named ERβcx for c-terminal exchange, diverged at exon 7 (Ogawa et al. Nucleic Acids Res. 1998; 26(15):3505-12). In this case, 61 amino acids of wild-type protein sequence were substituted with a unique sequence encoding 26 amino acids. In transcription assays, the relative expression of this novel receptor had profound effects on estrogen-induced trans-activation. The evidence suggested that these effects were mediated through a heterodimerization mechanism. This isoform was shown to preferentially form heterodimers with ERα and was demonstrated to behave in a dominant negative fashion against ERα transactivation in vitro in cotransfection studies. ERβcx was shown therefore to be a potential inhibitor of estrogen (E2) action through specific interaction with the ERα isoform (Nucleic Acids Res. 1998; 26(15):3505-12).
In summary, ongoing research suggests that there exists complexity in estrogen signaling pathways and ER variants may contribute to estrogen pharmacology differently.
In order to understand the mechanism of estrogen action and ERβ regulation of gene transcription, it is important to isolate and characterize novel subtypes, variants, and/or isoforms of the ER. Once the underlying ER subtype, variant or isoform responsible for a particular disease state or pathological condition is determined, one may have a more accurate means of prognostigating the estrogen receptor related disease outcome; one may use the presence or amount of expression of the novel polynucleotide of the present invention and/or the polypeptide encoded by such polynucleotide for diagnosing associated pathological conditions or a susceptibility to an associated pathological condition; one may accurately follow therapies, develop gene specific and isoform specific therapies influenced by ER, and/or may develop new pharmaceutical drug targets.
Thus, there exists a need to identify new variants and isoforms and their protein products for the therapeutic treatment of human diseases. The present invention satisfies this need and provides related advantages as well.